Abstract

DEP domain–containing 5 protein (DEPDC5) is a repressor of the recently recognized amino acid–sensing branch of the mTORC1 pathway. So far, its function in the brain remains largely unknown. Germline loss-of-function mutations in DEPDC5 have emerged as a major cause of familial refractory focal epilepsies, with case reports of sudden unexpected death in epilepsy (SUDEP). Remarkably, a fraction of patients also develop focal cortical dysplasia (FCD), a neurodevelopmental cortical malformation. We therefore hypothesized that a somatic second-hit mutation arising during brain development may support the focal nature of the dysplasia. Here, using postoperative human tissue, we provide the proof of concept that a biallelic 2-hit — brain somatic and germline — mutational mechanism in DEPDC5 causes focal epilepsy with FCD. We discovered a mutation gradient with a higher rate of mosaicism in the seizure-onset zone than in the surrounding epileptogenic zone. Furthermore, we demonstrate the causality of a Depdc5 brain mosaic inactivation using CRISPR-Cas9 editing and in utero electroporation in a mouse model recapitulating focal epilepsy with FCD and SUDEP-like events. We further unveil a key role of Depdc5 in shaping dendrite and spine morphology of excitatory neurons. This study reveals promising therapeutic avenues for treating drug-resistant focal epilepsies with mTORC1-targeting molecules.

Figure 1

(A) Family pedigree. The black square indicates the patient with FCD-associated epilepsy. Sanger sequence chromatograms show the germline nonsense variant (c.856C>T/p.Arg286*, black arrowheads) in the mother and child, and the somatic nonsense variant (c.865C>T/p.Gln289*, red arrowhead) that was only detected in the DNA extracted from the seizure-onset zone (SOZ), and not from the surrounding epileptogenic zone (EZ). (B) Integrative genomics viewer displays mutated bases in red in aligned reads: c.856C>T in blood (51%, 828 of 1,618 reads) and SOZ cortex DNA (48%, 748/1551 reads) and c.865C>T in SOZ cortex DNA only (10%, 148 of 1,505 reads). The 2 variants were systematically observed on different reads, indicating that they are in trans configuration. (C) Immunostaining against pS6 protein (red) and the neuronal marker NeuN (green) of a paraffin-embedded brain section from the patient. DAPI is shown in blue. White arrowheads indicate enlarged pS6+ neurons. Scale bar: 50 μm.